Cell cycle dependence of gene transfer by lipoplex polyplex and recombinant adenovirus
Cell cycle dependence of gene transfer by lipoplex polyplex and recombinant adenovirus
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DOI：
10.1038/sj.gt.3301102
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发表时间：
2000-03-01
期刊：
影响因子：
5.1
通讯作者：
Wagner, E
中科院分区：
文献类型：
--
作者：
Brunner, S;Sauer, T;Wagner, E
本研究的目的是探讨细胞周期对转染率的影响。采用逆流离心法洗脱，避免了细胞化学处理可能产生的副作用。用这种方法对细胞群按大小和密度进行划分，并用不同的非病毒方法(脂质体、TfpLys和TfPEI)、腺病毒增强型转染法(AVET系统)和重组腺病毒对离散细胞周期特异性群对应的部分进行转染。研究发现，转染率与细胞周期阶段密切相关。与表达水平最低的G1期细胞相比，在S或G2期进行转染时，多聚阳离子或脂类为基础的转染系统的细胞荧光素酶活性提高了30-500倍以上。相比之下，重组腺病毒没有观察到这种效果，因为重组腺病毒的变化只有四倍。我们的结果表明，有丝分裂活性不仅通过脂复合体，也通过复合体，而不是具有有效的核进入机制的病毒系统，提示接近M期的转染可能是通过核膜破裂来促进的。此外，对G1细胞的低转染率表明，沉积在G1细胞中的DNA复合体可能没有保留足够长的时间来利用有丝分裂效应，或者转基因细胞通过S时相受到抑制。
The aim of this study was to investigate the influence of cell cycle on transfection efficiency. Counterflow centrifugal elutriation was used which avoids possible side-effects from chemical treatment of cells. With this method, cell populations were fractionated by means of size and density, and fractions corresponding to discrete cell cycle phase-specific populations were transfected with various nonviral methods (Lipofectamine, TfpLys and TfPEI), adenovirus-enhanced transferrinfection (AVET system) and recombinant adenovirus. Transfection efficiency was found to be strongly dependent on the cell cycle stage at the time of transfection. Luciferase activity from cells transfected with polycation- or lipid-based transfection systems was 30- to more than 500-fold higher when transfection was performed during S or G2 phase compared with cells in G1 phase which have the lowest expression levels. In contrast, this effect was not observed with recombinant adenovirus which varied only four-fold. Our results indicate that mitotic activity enhances transfection not only by lipoplexes but also by polyplexes, but not a viral system which has an efficient nuclear entry machinery, suggesting that transfection close to M phase is facilitated perhaps by nuclear membrane breakdown. Furthermore, low transfection success into G1 cells indicates that DNA complexes deposited in G1 cells are probably not retained long enough to take advantage of mitosis effects or that passage of transfected cells through S phase is inhibitory.